Throughout this application various publications are referenced by the names of the authors and the year of publication within parentheses. Full citations for these publications may be found at the end of the specification immediately preceding the claims. The disclosures of these publications in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art to which this invention pertains.
Certain specific chromosomal translocations are known to be associated with human cancer (for a review see Yunis, 1983). The chromosomal rearrangements that occur as a result of these translocations may activate human cellular oncogenes that have been rearranged. Past studies of these cancers and the chromosomal translocations involved have focused on an analysis of the translocated oncogene sequences themselves.
This invention takes a novel approach in that it uses probes isolated from a limited region of the receptor chromosome which does not normally contain the deleterious gene. This limited region for which we propose the term "breakpoint cluster region" is a deoxyribonucleic acid molecule of limited length within which the various chromosomal breakpoints are clustered. Probes isolated from this region are used to detect a translocation by hybridizing the probes with chromosomal DNA and examining the resulting restriction patterns. Previous studies have identified breakpoints on the chromosomes involved in other cancers (Dalla-Favera et al., 1983; Adams et al., 1983; Erikson et al., 1983; Erikson et al. 1982). However, these breakpoints are distributed over a relatively large region of DNA and thus are not useful as hybridization probes to test for chromosomal translocations.
The breakpoint cluster region on human chromosome 22, which is involved in the formation of the Philadelphia chromosome, is the first such region to be discovered. It is contemplated that such regions exist on other human chromosomes and that this invention will be effective in detecting other diseases evidenced by chromosomal translocations.
Chronic myelocytic leukemia (CML) is characterized by the presence of the Philadelphia (Ph') chromosome in the leukemic cells of 96% of all CML patients. The Ph' chromosome is the result of a translocation between chromosome 22 and chromosome 9 (Rowley, 1973, 1982; Sanberg, 1980); its presence has important prognostic and diagnostic value. Previously we described the localization of the human c-abl oncogene (Heisterkamp et al., 1983a), to chromosome 9 (Heisterkamp et al., 1982) and demonstrated its translocation to the Philadelphia (22q.sup.-) chromosome in CML (de Klein et al., 1982). This demonstrated unequivocally that the t(9;22) is reciprocal. As the breakpoint on chromosome 9 is at the most telomeric band on this chromosome, q34, human c-abl must be translocated on a relatively small fragment (less than 5000 kb) to chromosome 22, suggesting a potential role for the c-abl oncogene in CML. This hypothesis was strengthened by the isolation of a chimeric DNA fragment from one CML patient containing sequences from chromosome 9 and 22 and located 14 kb immediately 5' of human v-abl homologous sequences (Heisterkamp et al., 1983b). We have used the chromosome 22-specific sequences of the chimeric DNA fragment to isolate a second chimeric chromosome 9/22 (9q.sup.+) fragment from a different CML patient. The chromosome 9-specific sequences in this fragment must be localized at a minimal distance of 40 kb from the human v-abl homologous sequences. Using the same probe, we have isolated an extended region on chromosome 22 from non-CML human DNA. In contrast to the situation on chromosome 9, the breakpoints on chromosome 22 in the DNAs of these two CML patients had occurred in the same region, although not at an identical site. We investigated the genomic organization of this area in a number of other Ph'-positive CML patients: all exhibited abnormal restriction enzyme patterns, indicating that in Ph-positive CML a breakpoint occurs in a single well defined region (less than 5.8 kb in length) of chromosome 22.